Our blog has been dormant for the best part of six months. That’s not because we haven’t been up to much though. In fact it’s been a very busy spell at Cryoconite Towers. We have been quietly working on samples and data from our last season on Svalbard – and preparing for this summer’s fieldwork. I thought the best way of offering a catchup is from the perspective of team member @icybear79

Who is @icybear79?

In a nutshell, icy bear is smarter than the average bear of little brain. Often thought to be Arwyn Edwards, Tris Irvine-Fynn or Sara Rassner (although they are some of “his people”), icy bear’s origins are shrouded in the mysterious north and will remain a secret as dark and chilling as a particularly filthy cryoconite hole. Icy bear first materialized at Aberystwyth as part of an exhibition on Climate Change last year.

It seems wherever ice is in crisis, icy bear is there.

It only seemed fair to bring icy bear with us to Svalbard last year as part our NERC project on glacier ecology

All too soon it was time for the Aber cryo team to return to the UK. It’s fair to say @icybear79 is at home in the field, but he was soon to become a telly academic in his own right

Appearing on daytime S4C…

…Outreach events…

…and scientific conferences, ranging from the International Glaciological Society’s British Branch, to the ASB Life in the Cold Workshop at Leeds, before reaching the dizzy heights of the Cambrian News

But of course – remember that wherever ice is in crisis, icy bear is there. By April it was high time for icy bear to head out again (having broken our lab’s ice machine)

Icy bear first headed South. Further South than any bear had been before.

Almost!

All the way south to South Island, New Zealand to help with teaching a Geography fieldcourse and looking at evidence of an outburst flood as part of a British Society of Geomorphology fieldwork grant.

As I write this, icybear79 is in Kathmandu in Nepal getting ready to head up to the Khumbu as part of a Royal Society project team looking at glaciers in the Himalaya. Hear the team’s leader, Dr. Ann Rowan, talk on BBC Radio 4 about the planned fieldwork. Icy bear (along with helper Tris Irvine-Fynn) will be looking at the hydrochemistry and maybe some microbiology while they’re there.

Tris will be taking icy bear to Arctic Canada next, where glaciers are changing rapidly, and will be working on a Climate Change Consortium of Wales project led by TrisTris Irvine-Fynn and Arwyn Edwards extracting a shallow ice core from a glacier. Expect more of this kind of thing!

Meanwhile, Arwyn will be heading to Svalbard for a week at UNIS. Unfortunately, they say you can never go home, and this is the case for Icy Bear too, so he’ll stay in Canada for the duration.

We (Icy Bear, Tris, Arwyn) will head straight from Greenland to join up with PhD students Ottavia & Stephen in Tarfala, in Northern Sweden as part of a EU FP7 InterAct project based there. After a few days’ respite, we then plan to head to finish off the season in the Alps.

After all of this, we may all be roundly sick of glaciers and fieldwork. But there will be no rest for the wicked, and Icy Bear won’t get any time off either. Pretty soon after all of this, Arwyn and Icy Bear will be heading to South Georgia as part of a collaboration with the Natural History Museum and National Geographic to examine its fragile glacial ecosystems. Finally, just today we received a request for Icy Bear’s very particular skill sets in Drønning Maud Land as part of a UK-US-Swedish project involving Aberystwyth’s Professor Neil Glasser in 2015-6.

Well, on Monday night I had great fun sharing my dirty secrets about microbes and glaciers with a lovely crew of people who turned up to the Aberystwyth Science Café.

Rather than rehash what I said, I thought it might be fun to report some of what the audience asked me at the end of the talk or at the bar. I was really humbled by the interest and pertinence of the questions, which came from a diverse and varied audience. I could recognize many eminent experts in palaeoglaciology, sedimentology, bioinformatics, mycology and botany in the audience, as well as a range of students and people just curious about all of this glacier bugs malarkey. So here’s what I could remember, as I remember it (a government health warning if there ever was!).

Radionuclides – from accidents and atmospheric nuclear tests. Alpine cryoconite is the most-enriched substance beyond test sites known to science. You can detect the radionuclide signatures of particular events even.

Mercury – Microbes in High Arctic snow have to deal with mercury contamination, and their plasmid genomes reflect this.

Persistent organic pollutants. Mainly on glaciers heavily used for skiing and recreation. Some cryoconite microbes seem very capable of bioremediating these pollutants.

2. Nature always tries to find a balance. Are the microbes in glaciers collaborating, competing etc?

Yes. Cryoconite, for instance, is a collaboration between phototrophs and heterotrophs. But there’s also lots of competition and bug-eat-bug stuff going on. In particular the viruses – these are highly abundant and their tendency to kill most of the bacteria but produce very few progeny – makes for one of the biggest viral shunts of the microbial loop known. Protozoal grazers are also important.

3. Is the effect of microbes on glaciers being accounted for in models of environmental change?

Surprisingly so. Rates of microbial activity in cryoconite (sat at 0.1-1 degrees C) have been compared to Mediterranean soils. We still lack really comprehensive, definitive audits of the cryoconite economy as it varies through space and time though.

6. Do they stay active if cryoconite is buried in the ice?

I don’t know. We’ve thought about this, and have some ideas from labwork about dormancy and resuscitation we’d love to test, (as an undergrad I studied in a lab which did a lot of this work) but we never made a sufficiently compelling case to be funded enough to find out in the real world.

7. The surface of one of Jupiter’s moons, Europa is water ice, and it’s coloured. Is that microbial pigments?

I honestly don’t know. It’s an interesting possibility, but it’s kind of beyond my logistical capabilities to organise a field trip there. I know there are many dedicated researchers working on astrobiology, and agencies which fund cold ecology as an analogue for extraterrestrial life, but I have to admit that I find more pressing concerns closer to home: The one planet we know supports rich and biodiverse life is experiencing warming which first affects the 80% of its biosphere which is colder than your fridge, and we have yet to find all of life on Earth, as we know it.

7. What does life on mean for primary succession in glacier forelands?

I’ll be spilling the beans about cryoconite and how the life of glaciers is surprisingly linked to microbial shenanigans at the Aberystwyth Science Café at 1930h on Monday the 11th of November at Aber Arts Centre. Plus there may be a guest appearance by @icybear79…

Bottom line up front: Snow covers ca. 30% of Earth’s surface; we are losing the last permanently dry snows in the Northern hemisphere: Are these vast (new) microbial habitats, thanks to climate change? We published a paper which shows bacteria can proliferate rapidly in a decaying Arctic snowpack.

You would be forgiven for thinking we’re all cryoconite – obsessed swivel-eyed loons here at AberCryoconite Towers. In reality, only some of us are. If only because we are also interested in other microbial habitats associated with glacial systems. In fact, if we can get our (numb, nitrile- gloved) hands on it, we’ll give anything icy a go. So, I’m going to summarize one of our recent papers, published byISME Journal on the dynamics of bacteria in High Arctic supraglacial snowpacks and released to the press last week.

Slush fund

Like the cryoconite metagenome paper, this paper was born out of a collaboration with the Innsbruck Crew supported by the Society for General Microbiology’s fund for research visits in 2010. Chatting with two of Professor Birgit Sattler’s grad students, Kathi Hell and Jakub Zarsky, it appeared that some of the molecular and statistical approaches I had been developing could complement their experimental fieldwork earlier in the summer, working on Larsbreen on Svalbard.

Larsbreen (left) and Longyearbreen

Kathi and Jakub had visited at a one-week interval, sampling in an up-glacier transect of three stations at kilometre intervals, digging three snowpits at each station to collect snow, slush and ice cores. The second visit had seen a thin layer of aeolian dust deposited on the snow surface, so they sampled that too. While Jakub measured the activity rates of the bacterial community, Kathi filtered the samples for DNA analysis and chemistry.

So, Kathi bought DNA extracts over to the Aber lab in March 2011 to do T-RFLP and pyrosequencing. This is what we found out:

So what is the story?

1. Welcome to the layer cake?

Essentially, profiling the bacterial community revealed differences between different layers of the snow-covered glacier. We think the particular differences could only be explained by changes in the bacterial population in situ, or in other words after deposition as postulated by Xiang et al. (2009). In particular, we found slush (i.e. snow, as it melts and goes horrible, mucky, slushy crap) to harbour a distinct bacterial community. This confirms the melting snowpack as an active habitat for bacteria.

2. Betaproteobacteria: Slush puppies?

Our amplicon pyrosequencing allowed us to track the fates of different taxonomic groups in the different layers between the two sampling days. We found that the major bacterial group (class) in our samples, Betaproteobacteria, remained consistently abundant, but the organisms in the group shifted around. In particular, the genus Polaromonas proved to be a smooth player, able to duck and dive, wheel and deal in the rapidly changing environs of the slush layer.

If I had to anthropomorphize the view I have of Polaromonas following this study, they would be supraglacial Del Boy type characters, playing it nice and cool, while all the other bacteria remain oblivious, just like Trigger. Polaromonas, as a genus, crops up fairly regularly in cold environments. You would therefore expect it to be something of a “Ronseal” type bug: does what it says on the tin. But members of the genus are expert in taking on dodgy deals elsewhere too: Polaromonas napthalenivorans, for instance loves to split horrible organic pollutants straight down the middle, sixty-forty. Because of this (and other, unpublished) work we are now taking a very close look at Polaromonas.

3. Pyrosequencing- the Aristotlean connnection.

When I first examined our amplicon reads, a good fraction were “unclassified bacteria” and I didn’t know what (if anything) to make of the data. Upon re-analysis, we found that these reads were in OTUs comprising members of Chlamydomonadaceae. Specifically, the reads aligned to plastid rRNAs; thanks to our primers struggling to remain specific to bacteria we gained some bonus data. Something of an Eureka! moment ensued: the reads’ taxonomy and physical distribution were consistent with snow algae.

Snow algae in a bag

Classicists might complain that I’ve got the wrong Greek: after all Eureka is apparently what Archimedes shouted, not Aristotle. But that isn’t the connection I was alluding to. Aristotle, amongst other things, was the first to observe snow algal blooms. These occur as green algae, Chlamydomonas nivalis, having grown within the snowpack, produce characteristic carotenoid-rich structures which can colour the top 10-20 cm of a snowpack blood red. It’s an unlikely spectacular. The first time I saw such a bloom with my own eyes it was several weeks into a trip in the Arctic. I thought someone had laid into the stash of rhodamine to create a practical joke on an epic scale…

Secondly, snow algal blooms demonstrate that snowpacks can be loci of considerable biological activity, resulting in spatially extensive phenomena which reveal themselves at brief timescales. Algal blooms are well known at sea too:

An algal bloom off the coast of England in 1999. Landsat image in the public domain (NASA)

4. Nitrogen pollution of glaciers is NO joke.

Serious chemists won’t like that subheading. Nevertheless, anthropogenic nitrogen pollution from faraway lands acts to fertilize nutrient-poor environments in the Arctic. Arctic glaciers and the Greenland ice sheet are no exception. Microbes in glacial ecosystems can respond to assimilate ammonium from even a single deposition event. Using 454, fingerprinting and qPCR methods we found specific populations of Betaproteobacteria associated with ammonia oxidation in the snowpack, and in particular on the ice surface. We also appeared to find correlations between the bacterial community revealed by 454 and the decoupling of nutrient and non-nutrient anion dynamics in the snowpack. This would suggest microbes in the snowpack environment are tucking in to the nitrogen pollution. Our observations add to the growing body of literature on microbial interactions with the molested nitrogen cycle of glaciers to reveal a picture of an unholy trinity between microbial processes, anthropogenic nitrogen pollution and unstable climate.

What does it all mean then?

I struggled (visibly, even in the final edition) to write the paper’s discussion in a coherent fashion. Fortunately one lunchtime in the café I bumped into Aberystwyth’s Greenland guru and BBC Frozen Planet / Operation Iceberg “duderino” (his words) Dr. Alun Hubbard. He took a few minutes out of revising a manuscript to a headline journal to describe conditions high on the Greenland Ice Sheet in July 2012. Slush was forming, and piling up, saturating the firn and running off. In just 4 days, the percentage of the Greenland ice sheet experiencing surface melting skyrocketed from ca. 40% to a maximum of 97% on the 12th of July. At that timescale, the melt is too “flashy” to let the algae get out the starting blocks. But our paper suggested the bacteria might.

Could it be that the epic melt on Greenland in 2012 triggered a massive bacterial bloom?

Who knows. From our Svalbard study we could see a heady mix of a globally ubiquitous genus of bacteria proliferating at similar timescales in similar conditions, interacting with nitrogen pollution to create a glacier-scale bacterial bloom. Plans to test this hypothesis have amounted to little thus far: 2013 saw far less melting, and attempts to reconstruct events from 2012 by looking at the firn from 2012 must take care as a bloom may not be conserved in the stratigraphy (according to our study).

Gee – I sure wish we had one of those doomsday machines – General Buck Turgidson

Previously, on 24, I offered up the back story of our new paper detailing a cryoconite metagenome. Writing it brought out the inarticulate cod philosopher in me with a tenuous analogy about “next-gen” sequencing and whether it is a disruptive technology or not. Rather than cut a short story long, and sour the rare note of optimism in seizing opportunities I thought a separate post was in order.

The emergence of “Next Gen” DNA sequencing, has, like the development of a new weapon, say The Bomb, proven truly explosive.

It has also proliferated widely, perhaps into hands which shouldn’t be trusted with it (like mine), and has sparked an arms race, if not a revolution.

The notion it has forced regime change in genomics is frequently encountered. A graph of cost per nucleotide vs time is an ubiquitous powerpoint offering to the extent of cliché.

This regime change has often been referred to as a democratization of genomics. It is a powerful and attractive notion: taking the power to sequence away from an industrial sequencing centre and into the hands of the “ordinary” researcher. Big science for the little guy; something like a physicist finding a Large Hadron Collider in her basement ten years from now.

Has that really happened though?

At the level of many individual researchers, buying in to a “next-gen” experiment is not cheap. Monetarily, it was certainly beyond my means as a brand-new independent researcher at the time of sequencing the cryoconite metagenome discussed earlier. I am grateful for colleagues willing to offer that opportunity rather than re-sequence standards. Otherwise, early steps with “Next Gen” can be tentative and fraught with failed runs. Within the scope of seedcorn and pump-priming funds one might wish to use for such high-risk exploration of a new methodology, it is very easy to burn your budget on a single failed experiment. If, however one is analysing samples from deep polar fieldwork, the “Next Gen” bit of an experimental framework can be the cheapest, especially if you have collected those samples using helicopters to access your field sites (£30 a minute and up). It becomes a matter of scale and perspective.

At the level of an individual institute, it can be a risky process too. As the fates and fortunes of various platforms play out, buying into a particular platform is a brave step. After an initial hurrah, and much longer to optimize it to gain useful data, our first “next-gen” sequencer now lies neglected, all but obsolete within two years of its purchase (EDIT: Actually, it is now obsolete). I believe the number of its paper outputs to date can be counted using the fingers of one hand. Despite my better judgement, I remain partial to the Ion Torrent PGM, even when MiSeq, Proton, and HiSeq are also in house. I suspect he cost per base of sequencing matters less than the cost per experiment for a microbial community study: Everything is extrapolative (sequencing depth – rant for another time?). A dinky #314v2 chip will do me (and my budget) fine, thanks. Yet I have to admit the Downfall of Ion Torrent video had me laughing and wincing in even measure.

Of course, mileage may vary. A few minutes on Web of Science tracking the rate of publication and citation of some prominent (predominantly Stateside) microbial ecologists reveals a sharp inflexion upwards round about the time their pyrosequencers lost the new car smell. Those able to surf the wave of “Next Gen” rather then drown under it have done well.

I would argue that the regime change offered by “next-gen” sequencing is not as democratizing as it is destabilizing. I suspect Professor Mark Pallen briefly offers a related argument within this excellent talk on the application of metagenomics to a field (clinical microbiology) where 19th century technology is still the norm. Is there a risk of a gulf between those who can and the rest? How accessible is the technology to the real next-gen: students, early career researchers and the like? How can standards be agreed and verified as progress evolves rapidly? Or the hysteresis between the months taken to publish and days to conduct expensive experiments which may be consequently overtaken by competitors?

Even if the gold rush has past in the eyes of some, the yield and scale of data churned out has transformed the way we could aspire to do microbial ecology in a few years. Can this focus on cracking out loads of data be at the expense of insight and impact? A triumph of technology over science? A microbial genome has become devalued from meriting fanfare in a Nature paper to a brief non peer-reviewed obituary in Genome Announcements within a decade or so.

To build on an aside from Pallen’s presentation again: If Hall’s predictions of rising cost-per-base are borne out, is there a Fahrenheit-454 measured in units of bp/$? At this point it becomes more economic to transform the reams of sequence data into knowledge (and hence biology) by the means of further experimentation and analysis rather than simply short read archive it and move on to the next sequencing project?

It might even be arguable that it is at that Fahrenheit-454 that “Next-Gen” would cease to become a disruptive technology, and perhaps enable us to look for truths in biology again.

Maybe that the regime change has yet to reach Fahrenheit-454 is reflected in the common usage of the terminology “next gen” sequencing. Nearly a decade has passed since the first description of 454 sequencing. I have purposefully used quotation marks in this post to highlight this uneasy Luddite-tinged limbo. “Now gen” sequencing still doesn’t seem right, even if 454 itself is about to become ex-gen but it could reflect the reality better – for some. Perhaps “high throughput” dodges the bullet, and this is what I prefer to use, without quotation marks.

Argh. Call it what you like. The bottom line is that, like proliferated Nukes, the genie is not going to go back into the bottle. “Next-gen” in it many forms here to stay. Yeehah.

In a sentence, we sequenced a lot of DNA ripped from Alpine cryoconite and found most of it to be from devious bacteria forced to steal carbon and nutrients in order to survive. Don’t take our word for it: the metagenome should be publically available on MG-RAST (#4491734.3) for you to re-analyse for your own purposes. We haven’t quite finished with it yet, though, but don’t let that stop you.

So, rather than précis the paper’s scientific context, what I wanted to write about here was a little about the paper’s backstory. A defining theme is encapsulated in the pretentious Sun Tzu quote at the start: seizing opportunities.

The paper has its roots in an email I sent the awesome International Woman of Science (who has an award to prove it), Professor Birgit Sattler in March of 2010. I had finished my PhD a few months earlier, and despite being jobless, grantless and paperless I wasn’t quite done with science. Having worked with Birgit on Svalbard, I suggested an Alpine reunion might be fun. Opportunity #1 presented itself: the Society for General Microbiology’s Presidential Fund for Research Visits. I applied for, and duly won my first grant. An interview with the SGM about my experience was published here. In a nutshell, if you are an UK based early career microbiologist, apply for it. The perspective, experience and CV points of getting a grant to go do science elsewhere you will gain are worth it.

Six months later, I found myself in the Austrian Alps padding around a glacier called Rotmoosferner, collecting cryoconite as I went, following mostly the SOPs I had developed on Svalbard. Opportunity #2 presented itself in the form of the fearsome Professor Andy Hodson. Andy had come out to play and had brought his fave toy, an infra-red gas analyser with which to estimate cryoconite net ecosystem productivity with him. Later, two Brit male scientists left unsupervised with beer and a device to measure gas composition led to its natural conclusions: My farts are twice as potent as The Hod’s, it takes 10 square metres of an Arctic glacier to make them carbon netural, and he has yet to forgive me for the trauma inflicted on his IRGA. I digress: NEP and radiometric analyses of productivity suggested the cryoconites were typically heterotrophic. This provided essential context for our DNA work.

Back in the UK, I extracted the DNAs and did some other analyses and archived the DNA for posterity in -80*C. The story goes cold then for a little over a year. In the meantime, Aberystwyth University was investing heavily in next generation sequencing kit and expertise with the support of the BBSRC and the Welsh Government. Opportunity #3 presented itself when Dr. Justin Pachebat phoned up: The maiden run of an Illumina HiScanSQ was happening in 36h. Did I have anything interesting to sequence?

An hour later, the DNAs from Rotmoosferner were thawed enough to be quantified fluorometrically. If pooled, we had enough to make a paired-end library for sequencing. Justin immediately set about making the library: shearing, end-repairing, ligating, amplifying, quantifying and qualifying the library like a man possessed. By 4AM and after takeout pizza, tons of tips and scalpel blades, we had a library. My role in this was marginal: I can just about work spin columns and courier stuff in ice boxes.

In the morning, I took the library off to the Translational Genomics Facility on our other campus, and left it in some capable hands. Good news and bad news followed: Of eight libraries loaded onto the sequencer, ours was the one that worked, no doubt due to Justin’s extensive experience in making Illumina libraries and some luck.

The bad news was, the sequencer had blown up halfway through the run.

I exaggerate, but I gather somewhere inside an Illumina sequencer, there’s an USB dongle which must stay in contact at all times with the rest of the machine. Like the iPhone port on an Ion Torrent, I don’t know why that feature seemed like a good idea at the time for the designers. It lost the plot, and some short circuits followed.

We could salvage what amounted to 7 Gbp of single-end reads. Fortunately, once the sequencer was fixed, Opportunity #4 knocked. Our library, by virtue of having worked, could be used to titrate loadings of the chip. It seemed more interesting that just re-sequencing a phage genome standard, so the sequencing people just did it.

We now had more data than I could make sense of. 27 billion base pairs of it. Nearly 10x of a human genome.

Fortunately, Opportunity #5 presented itself: a 152-core, 1.5TB RAM high performance computing cluster fresh out the box. Assembling the metagenome took some clever work on the part of an expert next-gen bioinformatician which let me do the easy stuff on a seven year old laptop from PC World.

And so the plan came together.

Certain caveats should be applied. Nevertheless, opportunities multiply as they are seized. We may have sequenced and assembled a lot of cryoconite DNA into one metagenome, but it dates from only one time and place. But we now have dozens of glacier metagenomes in the works at Aber.

The analysis pipelines are less than perfect, particularly when dealing with microbes from poorly-explored habitats such as this. Subglacial flyfishers beware! Thus, problematic novelty is a particular challenge. Dedicated efforts to sequence, annotate and experiment with genomes of key organisms are required.

I remember looking at a KEGG map (biochemist’s wallpaper showing the metabolic circuitry of a cell) annotated with metabolic pathways reconstructed from a oceanic metagenome and pondering if/when we would ever achieve something similar for cryoconite. Two years later we published one. But does the presence of contigs affiliated to various functional categories mean anything in practice? Downstream -omics can help provide answers, which is why we are working with colleagues expert in metabolomics and metatranscriptomics. It was particularly gratifying to read that the paper had captured the imagination of another young researcher interested in cryoconite, leading him to ponder about transcriptomics experiments with cryoconite.

It’s clear that next-gen sequencing offers many opportunities – and challenges. My next post will consider Opportunity #6: How did I Stop Worrying and Learn to Love Next-Gen?

It’s time the Aber crew broke our radio silence. We’re now back from the Arctic, and have recovered from the “green-out” that returning to civilized society to a welter of conference attendances, doctoral thesis examining and even media work can incur. Not to mention lawns in dire need of TLC.

Please watch this space, as over the next few weeks we’ll be posting more about what life and doing science at 79 degrees North is like, and some of our new papers too. For now though, we are just starting to sort through our bumper harvest of deep-frozen samples and data from this field season, so a lot to keep us busy with.

And I think one team member is missing the Arctic a little too much because he needs to “chill out” between TV appearances. But don’t tell @icybear79 that I said that.